FOOD SAFETY FROM THE FARM TO THE FORK: INTEGRATED STRATEGIES AND INNOVATIONS FOR SOUTH CAROLINA

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: TERMINATED
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1011066
• Project No.: SC-1700535
• Project Start Date: Oct 11, 2016
• Project End Date: Sep 30, 2020

Project Director
Greene, AN.

Recipient Organization
CLEMSON UNIVERSITY
(N/A)
CLEMSON,SC 29634

Performing Department
Animal & Veterinary Sciences

Non Technical Summary
A disease outbreak in the animal livestock industry or from food animal products can have serious consequences to the rendering industry, food animal industry, food processing industries and the consumer. After the recent outbreak of Salmonella in eggs, initially blame was erroneously assigned to the rendering industry. With the current level of media coverage and rapid methods of disseminating information and misinformation, an outbreak of even a low pathogenicity disease could receive widespread publicity. Upon such an outbreak, miscommunication and lack of solid data could lead to economic instability in the rendering industry and its agricultural markets. It is imperative that the rendering industry have conclusive evidence on the effectiveness of rendering and post-process storage conditions to destroy and/or prevent the growth of animal disease pathogens. Additionally, it is imperative that non-vertebrate test methods are developed to screen if any isolated Salmonella have potential to be pathogenic. Thorough understanding of bacterial transfer from surfaces is at the heart of much food safety and rapid detection methods for bacterial contamination are crucially needed in all aspects of food safety research and quality control. Maintaining food safety is an on-going commitment for food animal production. There are many aspects still unknown and food safety research to ensure foodborne outbreak prevention is timely and needed. Protecting the public from foodborne illness is of paramount importance. Understanding the impacts of environmental factors on survival of pathogens and the mechanisms of bacterial transfer will help reduce negative consequences and promote food safety within food preparation settings. This research is needed since food safety is a continuing problem especially with the emphasis on minimally-processing of food and the desire and need for extended food shelf life. Thousands of U.S. citizens die each year from foodborne agents, food borne illness costs billions of dollars annually in the U.S. and recalls of tainted food costs the food industry an additional billions of dollars. New food products and processes are needed to expand and improve the food industry in South Carolina. This research will attract and retain food industries in the region to improve the economic status of South Carolinians. Hurdle technology has been used in the food industry with great success. This research will study hurdle methods for use in the animal feed and animal livestock industries to ensure prevention of disease transmission to the animals and thus, potentially to animal food products and humans. Maintaining safe food production systems beginning at the farm level are essential to continued economic viability of agricultural producing regions in the state of South Carolina as well as the entire country. In animal product food safety, the source of bacterial contamination can begin on the farm in animal feed. The initial portion of this project will investigate animal feed safety with food safety in mind. The project will continue into following bacterial transfer in food settings and finally the use of biosensors for keeping foods safe. This project has potential to expand and improve the food industry in South Carolina. This research could attract and retain food industries in the region to improve the economic status of South Carolinians.

Animal Health Component

0%

Research Effort Categories

Basic

33%

Applied

33%

Developmental

34%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
712	4010	1100	100%

Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
4010 - Bacteria;

Field Of Science
1100 - Bacteriology;

Keywords

food safety

bacterial transfer

bacterial pathogenicity

biosensors

salmonella

Goals / Objectives
The major goals of this project involve maintaining product safety from the farm to the fork. In animal product food safety, the source of bacterial contamination can begin on the farm in animal feed. The initial portion of this project will investigate animal feed safety with food safety in mind. The project will continue into following bacterial transfer in food settings and finally the use of biosensors for keeping foods safe. The research will contribute to the ongoing efforts to improve food safety from the farm to the fork. By ensuring microbial safety of animal feed ingredients, the incidence of animal-borne disease entities should be reduced throughout the animal product food chain. This will not only work to help keep South Carolinians safe from foodborne illness, but also will provide stability in the food animal industries by better defining conditions which protect animal feed ingredients from contamination. Continuing to the human food side of this farm to fork model, this study will follow potential avenues for contamination and ways to prevent it so that South Carolina residents and others have lowered risk for foodborne illness.There four objectives for this project are:Using commercially available rendered meal samples, examine the relationship between water activity and moisture content and its impact on survival of Salmonella in animal feed ingredient products;Determine potential pathogenicity of Salmonella strains isolated from animal feed ingredients using C. elegans;Determine bacterial transfer and survival in food settings that impact food safety/quality;Investigate biosensors for food safety.

Project Methods
Objective #1Samples of pet-food grade and feed-grade poultry and beef meat and bone meals as well as feather meal and blood meal will be requested from rendering plants in the U.S. The initial water activity of each sample will be measured in duplicate in water activity sample cups using a dew-point water activity meter. Initial moisture content in each meal sample will be determined in duplicate by three separate drying methods.Each sample will be placed in the environmental chamber set at 50% humidity at 28°C (83°F) for 24 h. Additional samples of the same products will be stored under the same conditions for 48 h and 96 h. The experiment will be repeated at each 70% and 90% humidity levels at 28°C (83°F). The water activity of each of the samples will be measured after the appropriate time interval.Objective #2Four strains of pathogenic Salmonella will be obtained and grown overnight in tryptic soy broth. Caenorhabditidis elegans Strain Bristol (N2) will be maintained on nematode growth media and fed Escherichia coli OP50. Nematode populations will be synchronized at the egg stage; larvae will be allowed to grow to the L4 adult stage. Pathogenicity of any isolated Salmonella strains will be examined by feeding C. elegans nematodes each strain of Salmonella for 24 hrs before transferring back to plates containing their normal E. coli OP50 food. Nematodes will be observed to determine alive/dead status each day in triplicate trials. This project will investigate use of a biological system for screening potentially pathogenic strains of Salmonella.Objective #3A non-pathogenic Escherichia coli strain JM109 labeled with jellyfish green fluorescent protein will be used as the bacteria for transfer. The competent bacterial cells will be electroporated in a Gene Pulser II (Bio-Rad) with plasmid vector pGFPuv (ClonTech, Palo Alto, CA). Transformants will be selected from isolated colonies grown on Luria-Bertani agar (LB) plates containing 100g of ampicillin/mL. The resulting ampicillin-resistant transformants should emit bright green fluorescence under a handheld long wavelength UV light. The stability of GFP label in E. coli will be determined by streaking on trypticase soy agar (TSA) plates containing 100 g ampicillin/mL for several generations. The Escherichia coli JM 109 stock cultures will be held in a -80°C freezer in vials containing TSB supplemented with 20% (v/v) glycerol (Sigma, St. Louis, MO, USA). After thawing, 0.1 ml of culture will be transferred to 10 ml TSB containing 0.5% ampicillin in 2 loosely screw-capped tubes and then the tubes will be incubated for 16-18h at 37°C with vigorous shaking. A second transfer will be prepared from this first transfer culture by adding 0.1 ml from the first transfer tube to another fresh 10 ml TSB with 0.5% ampicillin, and again incubated for 16-18h at 37°C with shaking.After incubation, the cells will be harvested by centrifugatio, then the pellet resuspended in 10 mL of sterile peptone solution (0.1%) to obtain a population of approximately 6-7 log colony forming units (CFU)/ml. Initial cell populations will be verified by enumeration of the cells following surface plating in TSA containing 0.5% ampicillin and incubating at 37°C for 24 h. The approximate number of bacteria used per inoculation will be 106 CFU/ml.For hand transfer testing, subjects' hands will be washed with warm water and soap, dried and then 1 mL of the E. coli with plasmid vector pGFPuv inoculum was deposited in the center of the dominate hand. The E. coli will be spread onto both hands by rubbing hands together for 30 seconds than allowed to air dry for 30 seconds. Each subject will then touch the surface in question. This step is then repeated five times and then the bacteria recovered from the contact surfaces by placement into separate stomacher bags, each with 20 mL of sterile 0.1% peptone. The surface and peptone will be mixed for 30 seconds in the bag. Both the right and the left hands of each subject will be placed into the sterile stomacher bag with 20 mL of sterile 0.1% peptone. Hands will be massaged for 30 seconds with the peptone solution being swirled around the inoculated hand making contact with all fingers, palm, and back of the hand. Then the recovery rinsates will be serially diluted then plated in duplicate on TSA agar, and evenly spread on the agar surface. These plates will be incubated at 35°C for 24 h for enumeration.Objective #4The objective is to develop a comprehensive bacterial sensing solution for food processing plants. The sensing scheme will comprise a distinct configuration addressing specific ranges of bacterial sensitivity/specificity, together providing a complementary platform for alerting and identifying bacterial contaminations. All sensor configurations will be based upon the unique chromatic properties of PDA described above. Our strategy aims for an on-site bacterial sensor involving a "first-stage" trigger - the sensing assemblies - producing an alert on the presence of bacteria. Importantly, the trigger should not miss contamination, but it needs to minimize false positives to prevent needless application of more time-consuming and costly techniques for bacterial identification and quantification. Specifically, the tier 1 configurations will produce an alert through a visible color change, albeit without providing bacterial specificity.PDA swab detector Processed cotton is composed of ~99% cellulose, which, like alginate, provides a suitable matrix for immobilizing PDA assemblies. Our goal in this component of the research program will be to create "swab sensors" by immobilizing PDA onto cotton and other fibrous materials; these composite materials will subsequently be implemented for rapid detection of bacteria on exposed surfaces and similar locations in food processing plants.Our synthetic approach will be aided by the fact that carboxylic acid-terminated diacetylenes (such as PDA) are similar to amphiphillic surfactants used in the dyeing industry to enhance binding of dye molecules, and as such should bind quite well to cotton and other fibers. Indeed, we found that when cotton will be soaked in a diacetylene monomer solution and subsequently UV-irradiated, a stable blue PDA-dyed swab will be obtained. Furthermore, colorimetric transitions will be obtained by exposure of the swabs to bacterial populations in solutions and on surfaces. This configuration will be designed to identify bacterial biofilm formation, a critical pathogenic bacteria contaminant in food processing environments.The formation of thin PDA/sol-gel films comprising PDA domains embedded within a transparent sol-gel host matrix. The remarkable feature of the PDA/sol-gel films is the in situ response to bacterial biofilm formation through rapid colorimetric changes that can be detected by the naked eye. Importantly, the PDA/sol-gel patch constitutes a "passive" detector, and does not require any additional reagent addition and/or chemical reactions - in contrast to virtually all-current biofilm detection assays.PDA-functionalized filters. The bacteria from rinsate samples will be captured on polymer- or paper-based filters coupled to PDA. Cellulose acetate and nitrate filters will be the immediate focus of our studies, and techniques for tethering PDA to the filter matrix will be developed, similar to the approaches described above for the PDA-swab configuration. Monomer coupling through incubation/deposition will be particularly pursued, as it has shown promise in preliminary experiments. The filtering scheme could be particularly advantageous for concentrating bacterial samples and thereby enhancing detection sensitivity.

Progress 10/11/16 to 09/30/20

Outputs
Target Audience:The target audience of the water activity project and Salmonella project is the animal rendering industry. For food safety research we are targeting the general public and scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project allowed training of graduate students in use of water activity protocols and both undergraduate and graduate students to learn microbiological isolation and enumeration methods. How have the results been disseminated to communities of interest?The results were disseminated to the North American rendering industry through the Animal Co-Products Research & Education Center (ACREC) meetings. We have communicated food safety handling practices to the general public with live interviews and publications in the popular press. What do you plan to do during the next reporting period to accomplish the goals?We have concluded our research and will seek to publish the results.

Impacts
What was accomplished under these goals? Objective 1. Water Activity and Moisture Control Samples of pet-food grade and feed-grade poultry and beef meat and bone meals as well as feather meal and blood meal were received from rendering plants in the U.S. After exposure to 90% humidity levels at 28°C (83°F) for 96 h, the low initial water activity group for poultry meals increased to an average water activity level of 0.976±0.004. The medium initial water activity level group had reached an average water activity level of 0.979±0.005. The high initial water activity level group increased its average water activity level to 0.988±0.010. The water activity levels increased to a range in which Salmonella species can survive and potentially grow. The minimum reported aw for the growth of Salmonella species is 0.94. Exposure of rendered products to high humidity can lead to rapid increases in water activity of the products and can cause the products to develop favorable water activity conditions for microbial growth. Objective 2. Use of C. elegans as a sensor of Salmonella pathogenicity Salmonella enterica is a pathogenic, Gram negative bacterium which causes gastrointestinal infections in animals and humans resulting in diarrhea, abdominal cramps, vomiting and, in severe cases, even death. Salmonella isolates were screened for pathogenicity using Caenorhabditis elegans, a well-studied nematode, as a model. There is evidence that bacterial pathogens which cause infection in animals can also infect and reduce the lifespan of C. elegans significantly. Using known pathogenic Salmonella strains and isolated Salmonella strains, C. elegans (N2 wildtype) was tested for survival as compared to non-pathogenic bacterial controls using Escherichia coli OP50. Results indicated the pathogenic strains of Salmonella significantly reduced the lifespan of the C. elegans. Objective 3. Survival and transfer of bacteria on surfaces We continue to conduct research on food safety for the general public interest. During this period, we studied the sanitation of eating surfaces and transfer of bacteria during cleaning. Bacterial transfer is a concern when sharing food and drink, so to address this concern, this study examined the presence of microorganisms using the ATPase and the transfer of bacteria from one surface to others due to cleaning surfaces in sequence with the same cloth. Two experiments were performed to: 1) test random eating surfaces for the presence of microorganisms, and 2) transfer of bacteria from one surface to others by wiping 5 successive clean tile surfaces with the same cloth after the first surface had been inoculated with E. coli. In the first experiment, of the 165 randomly sampled eating surfaces, both 81% of the home eating surfaces and 81% of the public eating surfaces were categorized as unsanitary according to the ATPase testing. In the second experiment, 2.28 (cloth rag) - 3.4 logs (cotton gauze) were transferred from the first tile to the fifth tile by wiping each successive tile with a cloth initially clean before the first tile. This indicates that when wiping eating surfaces during cleaning with a cloth rag, other surfaces subsequently wiped will become contaminated. Objective 4. Microbial biosensors The purpose of this study was to develop a multi-layer polydiacetylene (PDA)-coated filters and to determined the proposed color response mechanism to bacteria. Unlike other reports which immobilize PDA liposomes on solid support, multi-layer PDA-coated filters were generated by directly evaporating organic solvents. Bacterial growth can trigger the color change of PDA sensors without any modification of PDA. The mechanism that pH change from bacteria metabolites lead to color change of PDA filters was verified by carefully designed dextrose-free medium with phenol red. Further, culturing Salmonella Typhimurium, E. coli, L. innocua and M. luteus with PDA-coated filters on phenol red agar in absence of dextrose verified the potential of applying PDA-coated filter for bacterial detection, specifically amine-producing bacteria. Thus, PDA-coated filters may be a useful tool for food safety and shelf life applications. Objective 5 - Anti-quality factors of water Water is considered to be the most important nutrient for plant, animal and human health and productivity. Recent droughts have increased the likelihood of degraded quality of water at water intakes for water treatment facilities as water levels drop and algal blooms become more prominent. In particular, the anti-quality factors, geosmin and 2-methylisoborneol (algal compounds) have been reported to increase with select algal blooms. In an effort to track the decrease in water quality as a result of drought and/or algal blooms, an ultra performance liquid chromatography (UPLC) separation of anti-quality factors (i.e., geosmin, 2-methylisoborneol [MIB]) coupled to a triple quad mass spectrometer (MS-MS) method (based on LC/MS-MS method of Bedner and Saito, 2020) has been adapted for use on our Waters Inc., UPLC MS/MS instrument (i.e., Acquity H Class coupled with TQ-S mass detector). Ionization of geosmin and MIB is accomplished with via Atmospheric Pressure Chemical Ionization in the positive mode. Our method separates and idenitifies geosmin and MIB in water samples in 2 minutes using a Waters Inc. 2.1 X 50 mm Acquity BEH C18 reverse phase column using an eluent gradient of water and methanol. At present, the method is preliminary and has a minimal detection limit of 35 ng on column for the unoptimized methodology. Detection limit will improve as the method is ionized. In addition, the method appears to be capable of separating and identifying enantiomers of geosmin and MIB, but will require further development and analysis fully delineate which peak is which enantiomer. It is unclear what, if any, differences exist between enantiomers of the compounds with regards to water quality. Having a method capable of quickly and accurately separating the enantiomers will enable future studies to define the potential differences. As currently developed, the method is suitable for qualitative assessment for presence of geosmin and MIB and relative quantification. However, additional development remains to improve detection limits, identify enantiomer peaks and finalize analytical grade quantification method.

Publications

• Type: Journal Articles Status: Accepted Year Published: 2020 Citation: Zhang, Y.* Tzeng, T-Z., Northcutt, J., Hanks, T., Pennington, W. and Dawson, P.L. 2020. A proposed mechanism to induce multi-layer polydiacetylene-coated filter color response to bacteria. Results in Chemistry. Accepted 7/2020. Published online.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Dawson, P.L. Hirt, D. Seydim, A*. 2020. Styrene monomer migration from expanded polystyrene into liquid food simulants. Journal of Food Research. 9 (3): 39-52.
• Type: Other Status: Published Year Published: 2020 Citation: Dawson, P.L. 4-20-2020. The House of Wellness TV (Australia) show. Zoom Interview on food safety during pandemic. Lauren Moore, producer M: [phone], E: [e-mail]
• Type: Other Status: Published Year Published: 2020 Citation: Dawson, P.L. 2-1-2019. Did You Just Eat That? NPR Ask Me Another, https://www.npr.org/2019/02/01/690661033/did-you-just-eat-that
• Type: Other Status: Published Year Published: 2020 Citation: Dawson, P.L. 6-20-2020. Australian Broadcasting Corporation. Jennifer McCuctheon. [e-mail] Article based on an interview and birthday candle study. https://www.abc.net.au/news/2020-06-20/blowing-out-the-candles-may-be-snuffed-out-thanks-to-coronavirus/12363056

Progress 10/01/18 to 09/30/19

Outputs
Target Audience:The target audience of the water activity project is the animal rendering industry. For food safety research, the general public and scientific community are the target audiences. The target audience for the animal production and health project would be animal producers and those specifically relatedto the safety of animal feeds (primarily mycotoxins). Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project allowed training of graduate students in use of water activity protocols and the inoculation and recovery of bacteria from microbial transfer studies as well as the training ofundergraduate students on the UPLC MS/MS instrument. How have the results been disseminated to communities of interest?The results were disseminated to the North American rendering industry through the Animal Co-Products Research & Education Center (ACREC) meetings. We have communicated food safety handling practices to the general public with live interviews and publications in the popular press. What do you plan to do during the next reporting period to accomplish the goals?We will continue our research and will seek to publish the results.

Impacts
What was accomplished under these goals? Objective #1 Water Activity and Moisture Content Temperature and humidity of the environmental chamber unit were monitored by a thermometer and humidity meter (90113-1, Springfield Precision, Oak Brook, IL). The humidity was controlled via a 1-gallon, drop humidifier (EE5301O, Crane USA, Bensenville, IL) connected to an electronic thermohygrostat controller with a humidity sensor (Plug and Play, Thermomart, Toronto, Canada). This unit allowed the humidity to be precisely controlled and monitored within the environmental chamber. Samples of poultry and beef meat and bone meals used for testing were randomly selected prior to experimentation. Each meat and bone meal sample was weighed (20 g) in duplicate into sterile, 50 mL Falcon tubes 89039-660, VWR Scientific Products, Suwanee, GA). The initial water activity was re-measured in two randomly selected meal samples prior to humidity exposure to determine if water activity changed while the meal samples were stored in 1-gallon, Ziploc® bags. The initial water activity of each sample was measured in duplicate in water activity sample cups (40107, Decagon Devices, Inc, Pullman, WA) using a dew-point water activity meter (Aqualab series 3TE, Decagon Devices, Inc., Pullman, WA). The water activity of pet-food grade and feed-grade poultry and beef meat and bone meals was measured after 24, 48 and 96 h time intervals of storage in 70% humidity levels. The purpose of the study was to determine if the meal samples absorbed moisture and if water activity levels were impacted. Results: After exposure to 70% humidity levels at 28°C (83°F) for 96 h, the low initial water activity group for poultry meals increased to an average water activity level of 0.737±0.017. The medium initial water activity level group had reached an average water activity level of 0.730±0.017. The high initial water activity level group increased its average water activity level to 0.714±0.026. After exposure to 70% humidity levels at 28°C (83°F) for 96 h, the low initial water activity group for beef meals rose to an average water activity level of 0.745±0.012. The medium initial water activity level group had reached an average water activity level of 0.734±0.005. The high initial water activity level group increased its average water activity level to 0.735±0.003. Objective 2: We continue to conduct research on food safety for the general public interest. During this period, we studied transfer of bacteria to cups and water during drinking. Bacterial transfer is a concern when sharing food and drink, so to address this concern, this study examined the bacterial transfer onto and into plastic drinking cups. Two experiments were performed to: 1) test bacterial transfer to the rim of a plastic drinking cup, and 2) test residual bacteria in the liquid that was consumed from the cup. Bacteria were enumerated from the cup rim and the water in the cup. The results for both types of transfer showed higher bacterial populations on cups exposed to drinking compared to those where no drinking occurred. However, there was more bacterial transfer to the rim of the plastic cup than the liquid inside the cup. The average number of bacteria recovered from the rim was in the 100,000 range compared to less than 50 organisms for the cups not contacted by the mouth during drinking. Similarly, the water from cups that had been sipped had over 1000 bacteria compared to less than 10 bacteria per 25 ml of water. Furthermore, the maximum number of bacteria recovered from the rim and water exposed to sipping was 1 million and 15,500, respectively. Several diseases (including the common cold, influenza, meningitis, rubella, chicken pox, measles, tuberculosis, cold sores and Staph infections) are spread from the mucus or skim of infected individuals. Since sharing of plastic drinking cups can contribute to the transfer of bacteria and other disease agents through both direct contact with the cup and by consumption of fluids containing transferred microbes, this practice should be limited or eliminated to minimize the risk of contracting disease. Objective 3: In this portion of the project, we intend to develop assays for measuring nutrients and secondary metabolites that may have beneficial or detrimental effects to animal production and health. Eventually we will partner with others to study the metabolism of selected compounds of interest (e.g., the mycotoxins like ergot alkaloids and algal compounds like geosmin). The UPLC MS/MS instrument has been repaired. A laboratory was obtained and altered to accommodate the UPLC MS/MS. The UPLC MS/MS has been moved to the new laboratory and is in working order, and we arenow starting to set up the laboratory. Started an undergraduate research group (4 undergrads + 3 grad students this past fall participated) - no data to report yet. We also started the process to develop hemp seed/oil secondary metabolite analysis and todevelop an assay for use in analyzing water samples for algal contaminating compounds (e.g., geosmin). We have recruited 12 undergraduates to work on these and related projects next semester.

Publications

• Type: Journal Articles Status: Published Year Published: 2018 Citation: Dawson, P.L. Hayley Thorson, Kelly Hooton, Justin Runey, David Hughes, Sally Foster, Melissa Roskosky, Sutton Fain-Swartz, Danielle Lynn, Inyee Han. 2018. Bacterial transfer to cups and water by drinking. Food and Nutrition Sciences. 9(12) 1386-1390.
• Type: Other Status: Published Year Published: 2018 Citation: Dawson, Paul. 5-30-2018. Live interview on Wisconsin Public Radio. Natalie Guyette Producer, Central Time on The Ideas Network 821 University Ave | Madison, WI 53706 [e-mail] | [phone] Follow @CentralTimeWPR on Twitter!
• Type: Other Status: Published Year Published: 2018 Citation: Jackson, Kate. 12-11-2018. Is airplane coffee or tea safe to drink. Today.com Article written from part of an interview. https://www.today.com/food/airplane-coffee-or-tea-safe-drink-t144818.
• Type: Other Status: Published Year Published: 2019 Citation: Dawson, P.L. 2-1-2019. Did You Just Eat That? NPR Ask Me Another, https://www.npr.org/2019/02/01/690661033/did-you-just-eat-that
• Type: Other Status: Published Year Published: 2019 Citation: Mayer, Johanna. 2019. Science Friday The origin of the five-second rule. February 20, 2019. https://www.sciencefriday.com/articles/the-origin-of-the-five-second-rule/
• Type: Other Status: Published Year Published: 2019 Citation: Dawson, P.L. July 16, 2019. Top of Mind with Julie Rose. BYU Radio. http://www.byuradio.org/episode/56423ddc-2630-4070-b8cf-e59b6ad3990b/top-of-mind-with-julie-rose-hurricanes-food-and-germs-rock-mollusks?playhead=1071&autoplay=true

Progress 10/01/17 to 09/30/18

Outputs
Target Audience:The target audience of the water activity project is the animal rendering industry. For food safety research we are targeting the general public and scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project allowed training of graduate students in use of water activity protocols and the inoculation and recovery of bacteria from microbial transfer studies. How have the results been disseminated to communities of interest?The results were disseminated to the North American rendering industry through the Animal Co-Products Research & Education Center (ACREC) meetings. We have communicated food safety handling practices to the general public with live interviews, publications in the popular press and journal articles. What do you plan to do during the next reporting period to accomplish the goals?We will continue our research that we are currently working on and will seek to publish the results.

Impacts
What was accomplished under these goals? Objective #1 Water Activity and Moisture Content Samples of pet-food grade and feed-grade poultry and beef meat and bone meals were supplied by rendering plants in the U.S. Twelve poultry by-product meal samples and twelve beef meal samples were randomly selected for this experiment. All samples were stored at room temperature in sealed, 1-gallon, Ziploc® bags until needed for experimentation. An environmental chamber was retrofitted to allow temperature and humidity modifications. Temperature and humidity of the environmental chamber unit were monitored by a thermometer and humidity meter (90113-1, Springfield Precision, Oak Brook, IL). The humidity was controlled via a 1-gallon humidifier (EE5301O, Crane USA, Bensenville, IL) connected to an electronic thermohygrostat controller with a humidity sensor (Plug and Play, Thermomart, Toronto, Canada). This unit allowed the humidity to be precisely controlled and monitored within the environmental chamber. Samples of poultry and beef meat and bone meals used for testing were randomly selected prior to experimentation. Each meat and bone meal sample was weighed (10 g) in duplicate into large 11x 2.9 cm aluminum pans (25433-022, VWR Scientific Products, Suwanee, GA). Each uncovered sample was placed in the environmental chamber (702-ASHR4, Labline Environette, Labline Instruments, Inc., Melrose Park, IL) set at 50% humidity at 28°C (83°F) for 24 h. Additional samples of the same products were stored under the same conditions for 48 h and 96 h. The experiment was repeated at each 70% and 90% humidity levels at 28°C (83°F). The water activity of each of the samples was measured after the appropriate time interval. The initial water activity was re-measured in two randomly selected meal samples prior to humidity exposure to determine if water activity changed while the meal samples were stored in 1-gallon, Ziploc® bags. All samples upon removal from the environmental chamber were immediately sealed in 1-gallon, Ziploc® bags until water activity measurements. For water activity measurement, samples were vigorously shaken and then transferred in duplicate (approximately 2-3 g) into water activity cups. The samples then were sealed in the water activity cups with parafilm (52858-032, VWR Scientific Products, Suwanee, GA and stored at room temperature in sealed, 1.5-gallon, Rubbermaid® storage container until water activity was measured and recorded (within 2 to 4 hours of sample collection). Results: Water activity levels of each poultry and beef meal sample increased with exposure to 50, 70, and 90% humidity levels at 28°C (83°F) when subsamples were measured at 24, 48, and 96 h. The poultry rendered meals were grouped together according to their initial water activity levels. The low initial water activity group had levels ranging from 0.204-0.239. The medium initial water activity level group had levels ranging from 0.246-0.268. The high initial water activity level group had levels ranging from 0.3515-0.501. After exposure to 50% humidity levels at 28°C (83°F) for 96 h, the low initial water activity group increased to an average water activity level of 0.552±0.006. The medium initial water activity level group had reached an average water activity level of 0.557±0.007. The high initial water activity level group increased its average water activity level to 0.560±0.015. The beef rendered meals were grouped together according to their initial water activity levels. The low initial water activity group had levels ranging from 0.2215-0.286. The medium initial water activity level group had levels ranging from 0.327-0.3405. The high initial water activity level group had levels ranging from 0.4285-0.474. Exposure to 50% humidity levels at 28°C (83°F) for 96 h, the low initial water activity group increased to an average water activity level of 0.558±0.009. The medium initial water activity level group had reached an average water activity level of 0.529±0.005. The high initial water activity level group increased its average water activity level to 0.571±0.004. Objective 2: Research for Objective 2 will commence upon completion of Objective 1. Objective 3: We continue to conduct research on food safety for the general public interest. During this period we studied transfer and survival of bacteria from flies to food. Two separate experiments were conducted to determine the transfer of E. coli by fruit flies during short term exposure to apple slices and bologna. No difference (P≤0.05) in the number of bacteria transferred to flies were found due to these exposure times. More bacteria were transferred to bologna at 1 and 5 min compared to apple while the number transferred did not differ at 15 min exposure. The percentage of E. coli transferred from inoculated food to flies was low (<0.5%) while the percentage transferred from flies to un-inoculated food was relatively high (>50%). This study found that flies can pick up and transfer bacteria to food in short exposure times. Objective 4: Research for Objective 4 will commence upon completion of Objective 3.

Publications

• Type: Books Status: Published Year Published: 2018 Citation: Dawson, P.L. and Sheldon, B.W. 2018. Did You Just Eat That? W.W. Norton and Company, 500 Fifth Avenue, New York, NY 10110
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Buyukyavuz, A., Benson, E., Song, J., Ellis, B., Han, I. and Dawson, P. 2018. Escherichia Coli transfer to food by fruit flies during short time exposure. Journal of Food Research. 7(4): 131-140 https://doi.org/10.5539/jfr.v7n4p131

Progress 10/11/16 to 09/30/17

Outputs
Target Audience:The target audience of the water activity project is the animal rendering industry. For food safety research we are targeting the general public and scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?This project allowed training of graduate students in use of water activity protocols and the inoculation and recovery of bacteria from microbial transfer studies. How have the results been disseminated to communities of interest?The results were disseminated to the North American rendering industry through the Animal Co-Products Research & Education Center (ACREC) meetings. We have communicated food safety handling practices to the general public with live interviews and publications in the popular press. What do you plan to do during the next reporting period to accomplish the goals?We will continue our research and will seek to publish the results.

Impacts
What was accomplished under these goals? Objective #1 Water Activity and Moisture Content Samples of pet-food grade and feed-grade poultry and beef meat and bone meals as well as feather meal and blood meal were received from rendering plants in the U.S. All samples were stored at room temperature in sealed, 1-gallon, Ziploc® bags until needed for experimentation. Water activity levels and moisture contents of pet-food grade and feed-grade poultry and beef meat and bone meals as well as feather meal and blood meals were measured to determine if there was a correlation between the two different types of measurements. Also, this experiment would determine if the water activity levels of the meals were within a survivable range for Salmonella species. The initial water activity of each sample was measured in duplicate in water activity sample cups (40107, Decagon Devices, Inc, Pullman, WA) using a dew-point water activity meter (Aqualab series 3TE, Decagon Devices, Inc., Pullman, WA). Initial moisture content in each meal sample was determined in duplicate by three separate drying methods as follows: infrared radiation drying using an infrared moisture analyzer (IR-35, Denver Instrument, Denver, CO, USA), forced convection oven (FD-53, Binder Inc., Bohemia, NY) drying at 135 C for 2 h (AOAC 930.15), and forced convection oven (FD-53, Binder Inc., Bohemia, NY) drying at 104 C for 3 h (Thiex and Van Erem, 1999). To determine moisture content via the infrared moisture analyzer (IR-35, Denver Instrument, Denver, CO, USA), 11.0 x 2.9 cm aluminum drying pans (25433-022, VWR Scientific Products, Suwanee, GA) containing approximately 2-3 g of each meal sample were placed into the instrument. The hood of the instrument was then lowered to automatically signal the beginning of the moisture analysis test. Moisture content (%) results for each meal sample were reported on the instrument's screen. To ascertain moisture content via the oven drying methods, a 5.7 x 1.6 cm aluminum drying pan with tab (25433-008, VWR Scientific Products, Suwanee, GA) was weighed (NewClassic ML4002E/03, Mettler Toledo International, Inc., Greifensee, Switzerland) and the tared weight (T) was recorded to the nearest 0.01 g. Each aluminum drying pan was tared to zero while on the balance and then 2 g of each well-mixed meal sample was added to each tared pan. The weight of each meal sample was recorded to the nearest 0.01g (M). Each meal sample was evenly distributed in the aluminum drying pan by gently shaking and then placed into a preheated oven (FD-53, Binder Inc., Bohemia, NY). The aluminum drying pans containing approximately 2 g of each meal sample were dried using each of the two oven drying methods: forced air oven drying at 135°C for 2 h (AOAC 930.15) and forced air oven drying at 104°C for 3 h (Thiex and Van Erem, 1999). Then aluminum drying pans containing the samples were allowed to cool in a dessicator. Once the dried samples cooled, the weight of the pan and dried samples was measured and recorded (S). The moisture content (%) of each sample was determined by Moisture Content (%) = 100-[(S-T)*100/M] Results: No correlations were apparent between moisture content measurements and water activity levels of the meals samples. The three moisture analysis methods produced very variable results. Objective 2: We continue to conduct research on food safety for the general public interest. During this period we studied transfer and survival of bacteria from hands to lemons and ice used for beverages, to salad bar tongs and when blowing birthday candles out. Three experiments were conducted for lemons and ice. In the first experiment, CFU per lemon and percentage of E. coli transferred were greater for wet lemons - 6123 cfu and 4.62% compared to 469 cfu and .2% for dry lemons. The second experiment found from 2 to 67% of the bacteria on hands were transferred to ice by hands and from 30 to 83% of the bacteria on scoops were transferred to ice. In a third experiment, lemons were inoculated with E. coli, then sliced and stored at 4 or 22C and tested at 0, 4 and 24 hr. Lemons stored at room temperature (22°C) had an increase in E. coli population after 24 hour while those stored under refrigeration had a decrease even though bacteria did survive on lemons in either case. For the salad tong study, the transfer of Escherichia coli from inoculated hands to salad bar tongs (experiment 1) and from inoculated tongs to hands (experiment 2) was determined in separate experiments. Transfer of E. coli averaged approximately 10% from hands to tongs and around 5% from tongs to hands. However, the transfer was as high as over 50% from both hands to tongs and tongs to hands. Handling of food bar tongs by multiple individuals could result in the transfer of bacteria and viruses between individuals and the spread of infectious agents. Blowing out candles over icing resulted in 15 times more and a statistically higher number of bacteria recovered from icing compared to icing that did not have candles blown out. Also, the variation (range) in bacteria recovered from icing was 100 times greater for icing exposed to the blow compared to the no blow treatment. Furthermore, the median and maximum transfer of bacteria increased 300 and 12,000%, respectively, due to blowing out candles

Publications

• Type: Other Status: Published Year Published: 2017 Citation: Zhang, [e-mail] 7/27/2017. Dawson Interview. Birthday Candle Bacteria.. https://www.theatlantic.com/health/archive/2017/07/birthday-candle-bacteria/534987/ The Atlantic
• Type: Journal Articles Status: Published Year Published: 2016 Citation: Bartz, M., Buyukyavuz, A., Dawson, E., Diener, M., Gates, R., Han, I., Johnson, L., Marvin, P., Musselwhite, C., Nicholson, A., Randar, D., Ritterpusch, M., Zazzara, M., Dawson, P. 2016. Transfer of Escherichia coli while using salad tongs. Journal of Food Microbiology, Safety and Hygiene. 1(2): 112 Open access journal.
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Dawson, P., Buyukyavuz, A., Aljeddawi, W. Martinez-Dawson, R., Downs, R. Mattox, C., Kurtz, A., MacInnis, M., Freeland, J., Garrison, S., May, T., McClary, J., Monito, F., Nguyen, T., Polte, K., Suffern, M., Tanner, Z., Thurmond, A. and Ellis, V. 2017. Transfer of Escherichia coli to lemon slices and ice during handling. Journal of Food Research 6 (5): 1-11
• Type: Journal Articles Status: Published Year Published: 2017 Citation: Dawson, P., Han, I., Lynn, D., Lackey, J., Baker, J. and Martinez-Dawson, R. 2017. Bacterial transfer associated with blowing out candles on a birthday cake. Journal of Food Research. 6(4): 1-5.
• Type: Other Status: Published Year Published: 2017 Citation: Haynes, Gavin, 3/`5/2017. Why science is obsessed with the five-second rule. The Guardian. https://www.theguardian.com/world/shortcuts/2017/mar/15/why-science-is-obsessed-with-the-five-second-rule?CMP=twt_gu
• Type: Other Status: Published Year Published: 2017 Citation: Hadley, Greg. 3/15/2017. The five second rule is actually real  at least sometimes  scientists say. Myrtle Beach Online. http://www.myrtlebeachonline.com/news/nation-world/world/article138716473.html
• Type: Other Status: Other Year Published: 2017 Citation: Miller, Abigail. 8/28/2017. Blowing out birthday candles increases bacteria 1400%, new study claims. Daily Mail Online Daily Mail Online |51 Astor Place, 9th floor, New York NY 10003 Tel: [phone] M: [phone] Email: [e-mail] http://www.dailymail.co.uk/health/article-4740590/Blowing-birthday-increases-bacteria-1400.html
• Type: Other Status: Published Year Published: 2017 Citation: Baker, Debbie. 8/31/2017. Study: Blowing out birthday candles makes for bacteria-frosted cakes. San Diego Tribune. http://www.sandiegouniontribune.com/news/whats-now/sd-me-birthday-cake-candles-20170731-story.html
• Type: Other Status: Published Year Published: 2017 Citation: Lillia Callum-Penso. 8/10/17. From an interview with Dawson; Caked with germs: How dirty is the cake when you blow out those candles? Greenville Online http://www.greenvilleonline.com/story/health/2017/08/08/caked-germs-how-dirty-cake-when-you-blow-out-those-candles/545139001/
• Type: Other Status: Published Year Published: 2017 Citation: Knight, Victoria. 8/11/2017, from Dawson interview CNN online article http://www.cnn.com/2017/08/10/health/science-dirty-food-habits-study/index.html [e-mail]
• Type: Other Status: Published Year Published: 2017 Citation: Dawson, Paul. 8/15/17 Appearance on the Poundstone Institute Podcast "Am I Wishing For a Germ Bath?" with Paula Poundstone and Adam Felber https://itunes.apple.com/us/podcast/live-from-the-poundstone-institute/ Ian Chillag Producer [e-mail]